Substance P signaling mediates BMP-dependent heterotopic ossification

Kan, Lixin; Lounev, Vitali; Pignolo, Robert J.; Duan, Lishu; Liu, Yijie; Stock, Stuart R.; McGuire, Tammy L.; Liu, Bao; Gerard, Norma P.; Shore, Eileen M.; Kaplan, Frederick S.; Kessler, John A.

doi:10.1002/jcb.23259

Cited by 99 publications

(133 citation statements)

References 72 publications

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“…Since these cells are behind the blood-nerve barrier [26], it is difficult to envision how BMP2 enters this location. However, the timing appears to be similar to induction of the neuro-inflammatory pathways previously described [12, 14]. The lack of PS + cells in the soft tissues outside the nerve may be a direct result of rapid binding of BMP2 by inhibitory binding proteins in the region [27].…”

Section: Discussionmentioning

confidence: 66%

“…One of the earliest steps in the process is the remodeling of the epineurial or outer matrix structure of the peripheral nerves [7, 12] [13] near the site of HO and associated neurogenic inflammation [12, 14]. This process involves mast cells, which degranulate and contribute to the activation of the sympathetic nervous system [12] ultimately leading to the formation of brown adipocytes from cells derived from the perineurial layer of the peripheral nerves [13].…”

Section: Introductionmentioning

confidence: 99%

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Progenitors in Peripheral Nerves Launch Heterotopic Ossification

Olmsted-Davis

Salisbury²,

Hoang³

et al. 2017

Stem Cells Translational Medicine

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Studies presented here, using a murine model of bone morphogenetic protein type 2 (BMP2)-induced HO show that the protein initiates HO by signaling through progenitors in the endoneurium of peripheral nerves. In the mouse, these cells were identified in the endoneurium one day after BMP2 induction using antibody against phosphoSMAD (PS) 1, 5, and 8. Studies conducted in a tracking mouse that contains a tamoxifen-regulated Wnt1-Cre recombinase crossed with a td Tomato red (TR) reporter (Wnt1CreErt:Ai9Tm) confirmed their neural origin. In this model both BMP2 induction and tamoxifen are absolutely required to induce TR. SP7+(osterix+)TR+ cells were found in the endoneurium on day 1 and associated with bone on day 7. Quantification of TR+ and TR− cells isolated by FACS showed that all SP7+ cells were found in the TR+ population, whereas only about 80 percent of the TR+ cells expressed SP7. Pre-chondrocytes (Sox 9+) and transient brown fat (tBAT, UCP1+) also co-expressed TR, suggesting that the progenitor in nerves is multi-potential. The endoneurium of human nerves near the site of HO contained many PS+ cells, and SP7+ cells were found in nerves and on bone in tissue from patients with HO. Control tissues and nerves did not contain these PS+ and SP7+ cells. Some osteoblasts on bone from patients with HO were positive for PS, suggesting the continued presence of BMP during bone formation. The data suggests that the progenitors for HO are derived from the endoneurium in both the mouse model of HO and in humans with HO.

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Section: Discussionmentioning

confidence: 66%

Section: Introductionmentioning

confidence: 99%

Progenitors in Peripheral Nerves Launch Heterotopic Ossification

Olmsted-Davis

Salisbury²,

Hoang³

et al. 2017

Stem Cells Translational Medicine

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“…Removal of the epineurium was correlated with migration of progenitors that reside in the perineurium that undergo brown adipogenesis [34], presumably for the purpose of patterning the new bone [27]. Blocking this process either through delivery of inhibitors of mast cell degranulation [33] or inhibitors of the binding of pain mediators to their receptor [13] resulted in a significant decrease in HO. Blocking nerve remodeling led to the accumulation within the endoneurium of nanog + Klf-4 + osterix + progenitors [33].…”

Section: Introductionmentioning

confidence: 99%

Osteoblasts Have a Neural Origin in Heterotopic Ossification

Lazard

Olmsted-Davis

Salisbury

et al. 2015

Clinical Orthopaedics &Amp; Related Research

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Background Heterotopic ossification (HO) is the process of bone formation at a nonskeletal site. Recently, we showed that the earliest steps occur in sensory nerves. We now extend these studies by identifying unique osteogenic progenitors within the endoneurial compartment of sensory nerves. Questions/purposes We asked: (1) What is the nature of the osteoprogenitor in the endoneurium of peripheral nerves? (2) How do osteoprogenitors travel from the nerve to the site of new bone formation? Methods HO was induced by intramuscular injection of Ad5BMP-2-transduced cells in mice. Osteoprogenitors were identified through immunohistochemistry and then quantified and further characterized by fluorescence-activated cell sorting and immunocytochemistry. The kinetics of the appearance of markers of extravasation was determined by quantitative reverse transcription-polymerase chain reaction. In each experiment mice were injected with bone morphogenetic protein-2 (BMP-2)-producing cells (experimental) or with cells transduced with empty vector or, in some cases, a group receiving no injection (control). Results Induction of HO leads to the expression, within 24 hours, of osteoblast-specific transcription factors in cells in the endoneurium followed by their coordinate disappearance from the nerve at 48 hours. They reappear in 123Clin Orthop Relat Res (2015) 473:2790-2806 DOI 10.1007 Clinical Orthopaedics and Related Research ® A Publication of The Association of Bone and Joint Surgeons® blood also at 48 hours after induction. During vessel entrance they begin to express the tight junction molecule, claudin 5. The cells expressing both the osteoblast-specific transcription factor, osterix, as well as claudin 5, then disappear from circulation at approximately 3 to 4 days by extravasation into the site of new bone formation. These endoneurial osteoprogenitors express neural markers PDGFRa, musashi-1, and the low-affinity nerve growth factor receptor p75(NTR) as well as the endothelial marker Tie-2. In a key experiment, cells that were obtained from mice that were injected with cells transduced with an empty vector, at 2 days after injection, contained 0.83% (SD, 0.07; 95% confidence interval [CI], 0.59-1.05) cells expressing claudin 5. However, cells that were obtained from mice 2 days after injection of BMP-2-producing cells contained 4.5% cells expressing claudin 5 (SD, 0.72%; 95% CI, 2.01-6.94; p \ 0.0015). Further analysis revealed that all of the cells expressing claudin 5 were found to be positive for osteoblast-specific markers, whereas cells not expressing claudin 5 were negative for these same markers. Conclusions The findings suggest that the endoneurial progenitors are the major osteogenic precursors that are used for HO. They exit the nerve through the endoneurial vessels, flow through vessels to the site of new bone formation, and then extravasate out of the vessels into this site. Clinical Relevance The biogenesis of osteoblasts in HO is very different than expected and shows that HO is, at least in par...

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“…It has been proposed that substance P might be one of systemic factors responsible for HO in mouse models of FOP [36,126]. Salisbury et al have shown that BMP-2 stimulation of substance P and calcitonin gene related peptide increased concentration of substance P in muscle [36].…”

Section: Sca1mentioning

confidence: 99%

“…Firstly, as studies on humans are retrospective, there is no study on the earliest stages of NHO to identify the early cellular and molecular events leading to heterotopic ossification in muscles and joints. Secondly, there is no unmodified animal model of NHO after central nervous system lesions [125,126]. The only existing animal models of HO are genetically modified mouse models of 1) progressive osseous heteroplasia, where HO is developed subcutaneously and caused by inactivating mutations of the GNAS gene [127], and 2) FOP in which activating mutations of ACVR1 are introduced as a transgene or knocked-in the endogenous mouse Acvr1 gene [48], or in which BMP-4 or BMP-2 are overexpressed by means of transgenes or recombinant adenoviruses [36,125,128].…”

Section: Introductionmentioning

confidence: 99%

Understanding heterotopic ossification after spinal cord injury

Kulina¹

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Neurological heterotopic ossification (NHO) is a frequent complication of spinal cord and traumatic brain injuries (15-25% of patients) and manifests as abnormal ossification of soft tissues near joints. NHO is very debilitating and further delays rehabilitation as it causes pain, joint deformation and ankylosis and vascular and nerve compression. In the absence of an animal model the mechanisms leading to NHO are unknown and consequently there is no preventive treatment. The only effective approach to eliminate HO is complicated and expensive surgical resection. However these surgeries are delicate and can be challenging as they are performed once the NHO are mature, often large and incapacitating entrapping large blood vessels and nerves.To elucidate NHO pathophysiology, we have developed the first animal model of NHO in genetically unmodified mice. Mice underwent a spinal cord transection (SCI); muscular inflammation was induced by intramuscular injection of cardiotoxin in limbs (IM-CTX).Formation of NHO was followed by μCT and immunohistology.SCI alone or muscular inflammation alone did not induce NHO in mice. The combination of both SCI and muscular inflammation was necessary to induce NHO. This is consistent with clinical observations as NHO incidence is higher in patients with severe trauma or concomitant infection. Abundant F4/80 + macrophages, which can provide pro-anabolic support in bone formation, were detected within the inflamed muscle and associated with areas of intramuscular bone formation (confirmed by von Kossa and collagen type 1 staining). In vivo depletion of phagocytic macrophages with clodronate-loaded liposomes prevented NHO formation whereas Zoledronate treatment exacerbated NHO. This supports our hypothesis that macrophage-mediated inflammation is a key activator of NHO following SCI. To further identify the source and type of macrophages, involved in the bone formation after SCI and test some potential treatment options different knock-out mouse models were investigated. My data suggest that local muscle residential macrophages potentially play an important role in NHO.In addition we identified several populations of muscle progenitor cells that are prone to osteogenic differentiation in vitro. These data suggest that the mesenchymal progenitors that differentiate into osteoblasts in HO following spinal cord injury are already present in healthy muscles and therefore can be derived from local muscle progenitor cells rather than being recruited from the remote site, such as bone marrow.iii Finally we investigated why SCI was necessary for NHO development. My hypothesis was that SCI causes release of systemic factors priming NHO. In support of this, I showed that NHO developed in non-paralysed inflamed front limbs of mice with SCI. Also, blood plasma from mice with SCI and IM-CTX induced osteogenic differentiation of cultured muscle mesenchymal progenitor cells (mMPC) sorted from naïve mice. This suggests that systemic factors facilitating NHO, such as substance P and G-CSF are ...

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Substance P signaling mediates BMP-dependent heterotopic ossification

Cited by 99 publications

References 72 publications

Progenitors in Peripheral Nerves Launch Heterotopic Ossification

Progenitors in Peripheral Nerves Launch Heterotopic Ossification

Osteoblasts Have a Neural Origin in Heterotopic Ossification

Understanding heterotopic ossification after spinal cord injury

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